Helper T lymphocytes play a central role in regulating immune responses. These cells specifically recognize antigen-derived peptides that are stably bound to class Il histocompatibility molecules (MHC) expressed on the surface of specialized antigen presenting cells (APC). The long-term objective of this project is to define the intracellular events and cofactors in APC that regulate the formation of peptide/MHC complexes. The first specific aim of the project is to determine how variation in the pH of intracellular compartments regulates peptide binding by mouse and human MHC molecules. Biochemical and mutational studies will be done to identify structural features of MHC that are responsible for this regulation. The 2nd specific aim is to identify other co-factors that facilitate peptide binding by MHC. Membrane reconstitution experiments will be used to define conditions necessary for efficient peptide binding by cell surface MHC and intracellular oligomers containing MHC and invariant chain. The 3d specific aim is to define mechanisms responsible for unfolding antigen and generation of the peptides that ultimately become associated with MHC. These mechanisms significantly influence the selection of antigenic determinants that become available for recognition by T cells. It is expected that the proposed study will broaden our understanding of critical events that are required for the initiation of protective immunity to infectious agents as well as deleterious responses to self antigens.